Abstract
Severe infection with the re-emerging enterovirus 71 (EV71 or EV-A71) can cause cardiopulmonary failure. However, in patients’ heart and lung, viral protein has not been detected. In mouse models, heart disease has not been reported. EV71-infected brainstem is generally believed to be responsible for the cardiopulmonary collapse. One major limitation in EV71 research is the lack of an efficient oral infection system using non-mouse-adapted clinical isolates. In a robust oral infection NOD/SCID mouse model, we detected EV71 protein at multiple organs, including heart and lung, in 100% of moribund mice with limb paralysis. Infiltrating leukocytes were always detected in heart and muscle, and VP1-positive M2 macrophages were abundant in the lung. Functional dissection on the pathogenesis mechanism revealed severe apoptosis, inflammatory cytokines, and abnormal electrocardiogram (EKG) in orally infected hearts. Therefore, cardiopulmonary disease could be one plausible cause of death in this mouse model. Inoculation of EV71 through an oral route resulted in viral infection in the intestine, viremia, and EV71 appeared to spread to peripheral tissues via blood circulation. Infectious virus was no longer detected in the blood on day 5 post-infection by the plaque formation assay. We demonstrated that both EV71 clinical isolate and cloned virus can target the cardiopulmonary system via a natural infection-like oral route.
Highlights
Epidemics of enterovirus 71 (EV71 or EV-A71) occurred frequently worldwide[1,2,3,4]
Because cardiopulmonary collapse is a fatal cause in severe cases of EV71 infected children, we examined the tissue distribution of EV71 in an oral infection mouse model (Fig. 1B)
EV71 specific RNA and plaque forming activity were extracted from dissected tissues (Materials and Methods), and measured by quantitative PCR (qPCR) and in vitro infection of RD cells, respectively (Fig. 1E–K)
Summary
Epidemics of enterovirus 71 (EV71 or EV-A71) occurred frequently worldwide[1,2,3,4]. In a recent outbreak in Shanghai, China, near 1000 deaths of children were reported[5]. Mouse-adapted strains accumulated many artificial mutations during serial passages through the mouse brain[20] They no longer can faithfully represent the parental clinical isolates from patients. Oral infection with clinical isolates of EV71 (106–107 pfu/mouse) was reported at a moderate efficiency around 15–40% in immunodeficient AG129 mice[22]. Efficient oral infection with EV71 clinical isolates (non-mouse-adapted) can be conducted only in the immunodeficient NOD/SCID mouse model. By using the more efficient oral infection system of NOD/SCID mice, we detected EV71 specific protein VP1, viral RNA, and infectivity by plaque forming assay in multiple organs, with near 100% frequencies of VP1 and viral RNA in heart and lung of moribund mice. The potential mechanisms of cardiomyocyte apoptosis and the route of virus spreading from gut to heart and lung are discussed
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